Experimental Study Of HUC-MSCs/hHGF Genetically Modified HUC-MSCs Transplantation On Repair Of Myocardial Injury

Objective:In this study, transplantation of human UC-MSCs (hUC-MSCs) cultured in vitro into infarct area was applied surgically on the Guangxi Bama miniswine model of AMI. The reparative effects of this therapy on ischemic myocardial injury were observed. Furthermore, whether a new strategy that combines hUC-MSCs transplantation and ex vivo human hepatocyte growth factor (hHGF) gene transferring with recombinant adenoviral vectors was more therapeutically efficient than hUC-MSCs cell therapy alone in the same AMI model was investigated, too.Methods:The hUC-MSCs isolated from human umbilical cord were cultured and expanded to passage5, and subsequently transfected by high titer adenoviral stocks expressing hHGF (Ad-HGF). The hUC-MSCs or hHGF genetically modified hUC-MSCs were labeled with CM-Dil before transplantation. The mid-third of LAD was ligated surgically to establish miniswine AMI model. The miniswines were randomly divided into four groups (n=6in each):(Group A) control group (without any treatment),(Group B) PBS group (PBS injection),(Group C) hUC-MSCs group (hUC-MSCs transplantation),(Group D) HGF-hUC-MSCs group (hHGF genetically modified hUC-MSCs transplantation). A total of2×107hUC-MSCs, HGF-hUC-MSCs or PBS of same volume were injected into the myocardial infarct area at9-10different points in the latter three groups. Six weeks later, cardiac perfusion and function were evaluated by SPECT and echocardiography in each group. The animals were euthanized and the tissues in infarct area were analyzed for the engraftment, proliferation and differentiation of stem cells, capillary density, viable myocardium, apoptosis and the expression of cytokines and growth factors to explore the underlying mechanisms.Results:(1) Six weeks later, the changes in MDP were significantly improved (P<0.001), EDWT and△WT%were increased significantly (P<0.001) in the hUC-MSCs and HGF-hUC-MSCs group compared with the control and PBS groups. Immunofluorescence results confirmed that the transplanted hUC-MSCs or HGF-hUC-MSCs were still alive and part of them appeared to have differentiated into cardiomyocytes and vascular endotheliums six weeks after transplantation. Meantime, it was also observed that resident cardiac stem cells (CSCs) differentiated into neonatal cardiomyocytes and vascular endotheliums, too. Angiogenesis was significantly enhanced by both hUC-MSCs group and HGF-hUC-MSCs group, which capillary density was higher than the control and PBS groups (P<0.001).The hUC-MSCs and HGF-hUC-MSCs groups showed more viable myocardial tissue in Masson’s trichrome staining (P<0.001), less cell number of apoptosis in the TUNEL analysis (P<0.001) than the control and PBS groups. The mRNA expression of cytokines and growth factors like VEGF、HGF、TGF-β3、vWF、SDF-1and CXCR4in local ischemic myocardium undergoing intramuscular cell transplantation was more abundant in hUC-MSCs and HGF-hUC-MSCs groups than the other two groups (P<0.001).(2) Six weeks after transplantion, there is no statistically difference in△MDP、EDWT and△WT%(P>0.05). However, the mRNA expression of HGF in local ischemic myocardium was up-regulated in HGF-hUC-MSCs group compared with hUC-MSCs group (P<0.05). It was also demonstrated that vWF staining analysis showed an enhanced angiogenesis (P=0.011) and Masson’s trichrome staining analysis showed an improvement in viable myocardium (P=0.007) in HGF-hUC-MSCs group relative to hUC-MSCs group.Conclusions:(1) hUC-MSCs and HGF-hUC-MSCs transplanted by direct injection into the infarct area could not only survived and differentiated, but also promoted CSCs recruitment and differentiation. The mechanisms may be involved exogenous and endogenous regeneration as well as paracrine action. In addition, hUC-MSCs and HGF-hUC-MSCs transplantation reduced apoptosis and fibrosis. enhanced viable myocardium and suppressed the ventricular remodeling, and thus improved myocardial perfusion and ventricular function.(2) The study shows that hHGF genetically modified hUC-MSCs transplantation therapy induced more potent angiogenesis and more viable myocardium than hUC-MSCs therapy alone. Stem cell-based angiogenic gene therapy may be a new therapeutic strategy for the treatment of severe ischemic cardiovascular disease.